Doxorubicin, an anthracycline antibiotic, is one of the most widely used and very potent anticancer agents. It has a broad spectrum of anticancer activity, being useful in the treatment of acute lymphoblastic leukemia, acute myeloblastic leukemia, Wilms' tumor, neuroblastoma, soft tissue and bone sarcomas, breast carcinoma, ovarian carcinoma, transitional cell bladder carcinoma, thyroid carcinoma, gastric carcinoma, Hodgkin's disease, malignant lymphoma, and bronchogenic carcinoma (doxorubicin package insert, Pfizer Inc., New York, N.Y.). In order to further exploit the enormous tumoricidal potential inherent in the structure of anthracycline antibiotics, thousands of synthetic derivatives have been described, including their analogs linked to various carrier macromolecules.
Nagy et al. (Proc. Natl. Acad. Sci. vol. 93, pp. 2464-2469, 1996) described that modifying doxorubicin (compound I) to 3′-deamino-3′-(2″-pyrroline-1″-yl)doxorubicin (compound II; 2-pyrrolinodoxorubicin) increased the anticancer potency towards human and mouse cancer cells in vitro 500 to 1000 times more than that of doxorubicin.

Doxorubicin rapidly enters the nucleus of cells and binds with high affinity to DNA by noncovalent intercalation between base pairs leading to inhibition of synthesis of biomacromolecules. It is generally accepted that biological effects of doxorubicin are associated with its ability to act as a topoisomerase II (topoII) poison perturbing the religation step of this enzyme and forming the ternary doxorubicin-DNA-topoII cleavable complex. 2-pyrrolino-doxorubicin appears to act in the same manner (Stepankova et al., Biochemical Pharmacology 82: 227-235, 2011). There are only small differences in DNA modifications by these anthracyclines and resulting conformational alterations in DNA. Similarly, the ability of 2-pyrrolinodoxorubicin modifications of DNA to inhibit catalytic activity of topoisomerase II does not differ significantly from that of doxorubicin. (Stepankova et al., supra)
However, increasing the potency of doxorubicin by converting it to a 2-pyrrolino compound rendered it nonspecific for cytotoxicity. Consequently, 2-pyrrolinodoxorubicin is toxic or lethal at or below doses that are required for an anticancer effect (Jungwirth, A et al., International Journal of Oncology 10: 877-884, 1997; and Szepeshazi, K et al., Clinical Cancer Research, 7: 2854-2861, 2001). As a result, 2-pyrrolinodoxorubicin has not been useful as a drug to treat cancer. Jungwirth et al., supra and Szepeshazi et al., supra have suggested modifying 2-pyrrolinodoxoubicin by conjugating it at the 14 position OH with somatostatin or luteinizing hormone-releasing hormone analogs. However, these modifications have made the resulting compounds specific for certain tumors, thereby reducing the broad spectrum activity of the parent compound.